ABSTRACT
The use of a small molecule compound to reduce toxic repeat RNA transcripts or their translated aberrant proteins to target repeat-expanded RNA/DNA with a G4C2 motif is a promising strategy to treat C9orf72-linked disorders. In this study, the crystal structures of DNA and RNA-DNA hybrid duplexes with the -GGGCCG- region as a G4C2 repeat motif were solved. Unusual groove widening and sharper bending of the G4C2 DNA duplex A-DNA conformation with B-form characteristics inside was observed. The G4C2 RNA-DNA hybrid duplex adopts a more typical rigid A form structure. Detailed structural analysis revealed that the G4C2 repeat motif of the DNA duplex exhibits a hydration shell and greater flexibility and serves as a 'hot-spot' for binding of the anthracene-based nickel complex, NiII(Chro)2 (Chro = Chromomycin A3). In addition to the original GGCC recognition site, NiII(Chro)2 has extended specificity and binds the flanked G:C base pairs of the GGCC core, resulting in minor groove contraction and straightening of the DNA backbone. We have also shown that Chro-metal complexes inhibit neuronal toxicity and suppresses locomotor deficits in a Drosophila model of C9orf72-associated ALS. The approach represents a new direction for drug discovery against ALS and FTD diseases by targeting G4C2 repeat motif DNA.
Subject(s)
Amyotrophic Lateral Sclerosis/drug therapy , C9orf72 Protein/genetics , DNA, A-Form/ultrastructure , Frontotemporal Dementia/drug therapy , Amyotrophic Lateral Sclerosis/genetics , Amyotrophic Lateral Sclerosis/pathology , Anthracenes/chemistry , Anthracenes/pharmacology , Coordination Complexes/chemistry , Coordination Complexes/pharmacology , DNA/drug effects , DNA/ultrastructure , DNA, A-Form/drug effects , Frontotemporal Dementia/genetics , Frontotemporal Dementia/pathology , Humans , Nucleic Acid Conformation/drug effects , Small Molecule Libraries/pharmacologyABSTRACT
We have reported the propensity of a DNA sequence containing CCG repeats to form a stable i-motif tetraplex structure in the absence of ligands. Here we show that an i-motif DNA sequence may transition to a base-extruded duplex structure with a GGCC tetranucleotide tract when bound to the (CoII)-mediated dimer of chromomycin A3, CoII(Chro)2. Biophysical experiments reveal that CCG trinucleotide repeats provide favorable binding sites for CoII(Chro)2. In addition, water hydration and divalent metal ion (CoII) interactions also play a crucial role in the stabilization of CCG trinucleotide repeats (TNRs). Our data furnish useful structural information for the design of novel therapeutic strategies to treat neurological diseases caused by repeat expansions.
Subject(s)
Chromomycin A3/pharmacology , Cobalt/pharmacology , Coordination Complexes/pharmacology , DNA/chemistry , Nucleic Acid Conformation/drug effects , Trinucleotide Repeats/drug effects , Chromomycin A3/chemistry , Cobalt/chemistry , Coordination Complexes/chemistry , Crystallography, X-Ray , Drug Discovery , Models, MolecularABSTRACT
Expansion of trinucleotide repeats (TNRs) within genes plays a major role in pathology of various neurological diseases. The correlations of these unusual repetitive sequences with the aetiology of these diseases and the mechanism by which those repeats are expanded during replication have been extensively studied. Small-molecule ligands that bind to TNRs could provide potent biological applications. First, the length of the TNR is the most important determinant of these neurological diseases. Ligands that reduce the repeat length or impair repeat expansion may be used to delay onset and reduce the severity of these diseases. Interestingly, many important anticancer ligands and antibiotics have desirable qualities when interacting with TNR DNA, and may form the basis for the development of novel therapeutics against neurological diseases. Second, designed ligands that bind to expanded TNRs with high specificity based on the structural and chemical characteristics of these repeats can serve as diagnostic tools for determining repeat length and may have applications in preventive medicine. In this article we will review our current understanding of the interaction between DNA-binding ligands and TNRs.
Subject(s)
DNA/chemistry , DNA/drug effects , Nervous System Diseases/diagnosis , Nervous System Diseases/drug therapy , Small Molecule Libraries/chemistry , Small Molecule Libraries/pharmacology , Trinucleotide Repeats/drug effects , Humans , Ligands , Nervous System Diseases/genetics , Trinucleotide Repeats/geneticsABSTRACT
CCG triplet repeats can fold into tetraplex structures, which are associated with the expansion of (CCG)n trinucleotide sequences in certain neurological diseases. These structures are stabilized by intertwining i-motifs. However, the structural basis for tetraplex i-motif formation in CCG triplet repeats remains largely unknown. We report the first crystal structure of a CCG-repeat sequence, which shows that two dT(CCG)3 A strands can associate to form a tetraplex structure with an i-motif core containing four C:C(+) pairs flanked by two G:G homopurine base pairs as a structural motif. The tetraplex core is attached to a short parallel-stranded duplex. Each hairpin itself contains a central CCG loop in which the nucleotides are flipped out and stabilized by stacking interactions. The helical twists between adjacent cytosine residues of this structure in the i-motif core have an average value of 30°, which is greater than those previously reported for i-motif structures.